Abstract
The reversible phosphorylation of proteins on serine, threonine, and tyrosine residues is one of the most important post-translational modifications that regulates many biological processes. The phosphoproteome has not been studied in any great detail in recombinant Chinese hamster ovary (CHO) cells to date despite phosphorylation playing a crucial role in regulating many molecular and cellular processes relevant to bioprocess phenotypes including, for example, transcription, translation, growth, apoptosis, and signal transduction. In this chapter, we provide a protocol for the phosphoproteomic analysis of Chinese hamster ovary cells using phosphopeptide enrichment with metal oxide affinity chromatography (MOAC) and immobilized metal affinity chromatography (IMAC) techniques, followed by site-specific identification of phosphorylated residues using LC-MS (MS2 and MS3) strategies.
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Acknowledgments
The authors acknowledge funding from Science Foundation Ireland (grant ref. 13/1A/1841) and the Horizon 2020 Marie Curie ITN programme – eCHO systems (grant ref.: 642663).
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Henry, M., Coleman, O., Prashant, Clynes, M., Meleady, P. (2017). Phosphopeptide Enrichment and LC-MS/MS Analysis to Study the Phosphoproteome of Recombinant Chinese Hamster Ovary Cells. In: Meleady, P. (eds) Heterologous Protein Production in CHO Cells. Methods in Molecular Biology, vol 1603. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-6972-2_13
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DOI: https://doi.org/10.1007/978-1-4939-6972-2_13
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